HIV/AIDS continues to be a major threat to human health. The use of combinations of small molecule drugs in highly active any-retoviral therapy (HAART) to stop or thwart HIV propagation has had a major impact on delaying the progression from HIV-1 infection to the development of AIDS. Despite this progress, there are problems associated with a lifetime of anti-viral small drug therapy which include toxicity, the emergence of virus resistant to multiple drugs, and the cost of a daily, lifelong medication. The proposed studies take advantage of recent advances in the functionality of sequence specific Zn finger and homing endonucleases are capable of destroying gene function. The endonucleases are being developed in other projects of this program which will target both the chemokine co-receptor for HIV CCR5 as well as integrated proviral regulatory regions. The major challenge in using these endonucleases in a therapeutic setting is delivery of the enzymes or sequences encoding the enzymes to HIV-1 infectible cells. Since long term expression of the endonucleases could lead to secondary non-target cleavage and gene destruction, transient expression is a necessity if this approach is to be clinically useful. The proposed studies take advantage of two novel platforms for delivery of Zn finger and homing endonuclease encoding sequences into HIV (and SHIV) infectible and infected cells. The first approach uses RNA aptamers with proven ability to bind to the HIV-1 envelope expressed on the surface of HUV infected cells and internalize for functional delivery of attached siRNAs. We now will test the ability of these aptamers along with an internalizing CD4 specific aptamer to deliver backbone modified, nuclease resistant mRNAs encoding the endocucleases to uninfected and SHIV infected cells. The targets for the nucleases are the CCR5 co-receptor gene and the SHIV LTRs. As an alternative delivery approach we will test flexible TAMAM dendrimers which we have demonstrated as effective agents for siRNA deliverty in cell culture and in vivo. We will utilize this non-toxic but non-targeted delivery platform as an alternative approach for delivering backbone modified mRNAs and/or cDNAs encoding the anti-viral and anti-CCR5 endonucleases. Since both approaches have been successfully tested in vivo in a humanized mouse model for HIV infection, we will proceed to develop these agents for future application in the SHIV-pigtail macaque model utilized in other projects of this proposal. Knowledge gained from the proposed studies will result in new in vivo approaches for delivery of DNA targeting endonucleases for the treatment, and potential purging of HIV-1 infected.